featurecounts and DESEQ
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2.9 years ago
anjuraas ▴ 10

hi I aligned my sequence with bowtie2 and put the BAM file into featurecount. And later the count table into deseq, but it is showing 'all samples have 0 counts for all genes'. Why my samples don't have miRNA?i downloaded the dataset from SRA database. please help

bowtie2 miRNA featurecount deseq • 1.5k views
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can you post a head of your count table?

and your featureCount command

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featurecount output

I am using usegalaxy.

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did you get any runtime log from it? are you using the correct gff (annotation) files?

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Please confirm that the chromosome identifiers match between your gff and alignment.

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yes. i checked that. Gene identifier ID is 'ID'. But its still showing 0 counts. Is there a chance that my read doesn't have miRNA?

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What WouterDeCoster means is : are the same IDs present in all your files? eg. in GFF file it says chr0 and in your mapping Chr00 or such?

Can you post a sample of the GFF you are using and of the genomic sequence (mainly the headers) you're mapping against?

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Indeed, and more commonly, for human genetics, for example GFF: chr1, alignment/fasta: 1.

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Commercial Photography this is my GFF file

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This is my bowtie sam file

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So your GFF contains NC_000001.11 and your sam file contains chr17... QED.

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what can I do?please help

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Use another gff which matches to the fasta you used for the alignment.

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hi, I used the in built gff file,and now its showing few counts.but still I cant get the differential expression.this is the error report I got from deseq.i am sorry, I cant understand its meaning.

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![Deseq error report][1]

Commercial Photography

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Which organism are you working on? How was the data generated? Did you get appropriate alignment metrics?

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I am working on dataset on gastric cancer downloaded from SRA database.its trueseq small RNA library using illumine Miseq. I aligned with bowtie 2 and got alignment rate above 90%.

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